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Functional and structural characterization of Hyp730, a highly conserved and dormancy‐specific hypothetical membrane protein

Membrane proteins represent major drug targets, and the ability to determine their functions, structures, and conformational changes will significantly advance mechanistic approaches to both biotechnology and bioremediation, as well as the fight against pathogenic bacteria. A pertinent example is My...

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Autores principales: Fannin, Stewart, Rangel, Jonathan, Bodurin, Abiodun P., Yu, Tannon, Mistretta, Brandon, Mali, Sujina, Gunaratne, Preethi, Bark, Steven J., Ebalunode, Jerry O., Khan, Arshad, Widger, William R., Sen, Mehmet
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7856521/
https://www.ncbi.nlm.nih.gov/pubmed/33650800
http://dx.doi.org/10.1002/mbo3.1154
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author Fannin, Stewart
Rangel, Jonathan
Bodurin, Abiodun P.
Yu, Tannon
Mistretta, Brandon
Mali, Sujina
Gunaratne, Preethi
Bark, Steven J.
Ebalunode, Jerry O.
Khan, Arshad
Widger, William R.
Sen, Mehmet
author_facet Fannin, Stewart
Rangel, Jonathan
Bodurin, Abiodun P.
Yu, Tannon
Mistretta, Brandon
Mali, Sujina
Gunaratne, Preethi
Bark, Steven J.
Ebalunode, Jerry O.
Khan, Arshad
Widger, William R.
Sen, Mehmet
author_sort Fannin, Stewart
collection PubMed
description Membrane proteins represent major drug targets, and the ability to determine their functions, structures, and conformational changes will significantly advance mechanistic approaches to both biotechnology and bioremediation, as well as the fight against pathogenic bacteria. A pertinent example is Mycobacterium tuberculosis (H37Rv), which contains ~4000 protein‐coding genes, with almost a thousand having been categorized as ‘membrane protein’, and a few of which (~1%) have been functionally characterized and structurally modeled. However, the functions and structures of most membrane proteins that are sparsely, or only transiently, expressed, but essential in small phenotypic subpopulations or under stress conditions such as persistence or dormancy, remain unknown. Our deep quantitative proteomics profiles revealed that the hypothetical membrane protein 730 (Hyp730) WP_010079730 (protein ID Mlut_RS11895) from M. luteus is upregulated in dormancy despite a ~5‐fold reduction in overall protein diversity. Its H37Rv paralog, Rv1234, showed a similar proteomic signature, but the function of Hyp730‐like proteins has never been characterized. Here, we present an extensive proteomic and transcriptomic analysis of Hyp730 and have also characterized its in vitro recombinant expression, purification, refolding, and essentiality as well as its tertiary fold. Our biophysical studies, circular dichroism, and tryptophan fluorescence are in immediate agreement with in‐depth in silico 3D‐structure prediction, suggesting that Hyp730 is a double‐pass membrane‐spanning protein. Ablation of Hyp730‐expression did not alter M. luteus growth, indicating that Hyp730 is not essential. Structural homology comparisons showed that Hyp730 is highly conserved and non‐redundant in G+C rich Actinobacteria and might be involved, under stress conditions, in an energy‐saving role in respiration during dormancy.
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spelling pubmed-78565212021-02-05 Functional and structural characterization of Hyp730, a highly conserved and dormancy‐specific hypothetical membrane protein Fannin, Stewart Rangel, Jonathan Bodurin, Abiodun P. Yu, Tannon Mistretta, Brandon Mali, Sujina Gunaratne, Preethi Bark, Steven J. Ebalunode, Jerry O. Khan, Arshad Widger, William R. Sen, Mehmet Microbiologyopen Original Articles Membrane proteins represent major drug targets, and the ability to determine their functions, structures, and conformational changes will significantly advance mechanistic approaches to both biotechnology and bioremediation, as well as the fight against pathogenic bacteria. A pertinent example is Mycobacterium tuberculosis (H37Rv), which contains ~4000 protein‐coding genes, with almost a thousand having been categorized as ‘membrane protein’, and a few of which (~1%) have been functionally characterized and structurally modeled. However, the functions and structures of most membrane proteins that are sparsely, or only transiently, expressed, but essential in small phenotypic subpopulations or under stress conditions such as persistence or dormancy, remain unknown. Our deep quantitative proteomics profiles revealed that the hypothetical membrane protein 730 (Hyp730) WP_010079730 (protein ID Mlut_RS11895) from M. luteus is upregulated in dormancy despite a ~5‐fold reduction in overall protein diversity. Its H37Rv paralog, Rv1234, showed a similar proteomic signature, but the function of Hyp730‐like proteins has never been characterized. Here, we present an extensive proteomic and transcriptomic analysis of Hyp730 and have also characterized its in vitro recombinant expression, purification, refolding, and essentiality as well as its tertiary fold. Our biophysical studies, circular dichroism, and tryptophan fluorescence are in immediate agreement with in‐depth in silico 3D‐structure prediction, suggesting that Hyp730 is a double‐pass membrane‐spanning protein. Ablation of Hyp730‐expression did not alter M. luteus growth, indicating that Hyp730 is not essential. Structural homology comparisons showed that Hyp730 is highly conserved and non‐redundant in G+C rich Actinobacteria and might be involved, under stress conditions, in an energy‐saving role in respiration during dormancy. John Wiley and Sons Inc. 2021-02-03 /pmc/articles/PMC7856521/ /pubmed/33650800 http://dx.doi.org/10.1002/mbo3.1154 Text en © 2021 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd. This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Original Articles
Fannin, Stewart
Rangel, Jonathan
Bodurin, Abiodun P.
Yu, Tannon
Mistretta, Brandon
Mali, Sujina
Gunaratne, Preethi
Bark, Steven J.
Ebalunode, Jerry O.
Khan, Arshad
Widger, William R.
Sen, Mehmet
Functional and structural characterization of Hyp730, a highly conserved and dormancy‐specific hypothetical membrane protein
title Functional and structural characterization of Hyp730, a highly conserved and dormancy‐specific hypothetical membrane protein
title_full Functional and structural characterization of Hyp730, a highly conserved and dormancy‐specific hypothetical membrane protein
title_fullStr Functional and structural characterization of Hyp730, a highly conserved and dormancy‐specific hypothetical membrane protein
title_full_unstemmed Functional and structural characterization of Hyp730, a highly conserved and dormancy‐specific hypothetical membrane protein
title_short Functional and structural characterization of Hyp730, a highly conserved and dormancy‐specific hypothetical membrane protein
title_sort functional and structural characterization of hyp730, a highly conserved and dormancy‐specific hypothetical membrane protein
topic Original Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7856521/
https://www.ncbi.nlm.nih.gov/pubmed/33650800
http://dx.doi.org/10.1002/mbo3.1154
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